10 Pseudobulk
Run reintegration after cleaning
10.1 REINTEGRATION
The integrated scRNAseq dataset of all brain cell populations was subsetted to retain those cells no subsetted out by the “expert committee”.
10.2 PCA
PC details
PC1 captures a strong myelination axis, with high loadings for oligodendrocyte and myelin-related genes (Plp1, Mbp, Mag, Mal, Car2, Cryab) and opposite contributions from astrocytic (Ptgds, Enpp2, Edil3) and microglial (Hexb, C1qa, Ctss, Cst3) markers.
PC2 reflects a neuron–astrocyte interaction gradient, driven by the astrocytic ECM regulator Sparcl1 on one pole and the neuronal signaling gene Plcb1 on the other.
PC3 highlights synaptic connectivity and neurotransmission, with neuronal adhesion and plasticity genes (Nrxn1, Ntm, Lsamp, Ctnnd2, Adgrb3, Ntrk2) and the inhibitory receptor Gabrb1 loading strongly, balanced by astrocytic transporters (Slc1a2, Slc4a4) and Ptprz1. Together, these axes describe complementary layers of variance corresponding to myelination, glial–neuronal signaling, and synaptic specialization.
10.3 UMAP after reintegration
| Adu | IgG | p.overall | |
|---|---|---|---|
| N=3 | N=3 | ||
| Astrocytes | 5.90 [3.90;8.90] | 7.70 [5.95;8.05] | 0.827 |
| BAM | 0.90 [0.85;0.90] | 0.40 [0.40;0.65] | 0.239 |
| Choroid | 2.30 [1.35;2.50] | 1.60 [0.90;2.70] | 0.827 |
| Endothelial | 7.20 [7.20;7.45] | 7.60 [7.30;9.65] | 0.825 |
| Ependymal | 0.30 [0.25;0.45] | 0.60 [0.45;0.65] | 0.261 |
| Microglia | 58.9 [56.3;62.3] | 59.1 [57.0;59.5] | 0.827 |
| NPC | 0.70 [0.55;0.85] | 1.60 [1.45;1.70] | 0.050 |
| Neurons | 1.10 [0.85;1.70] | 1.10 [0.85;1.85] | 0.822 |
| OPC | 0.30 [0.25;0.30] | 0.60 [0.40;0.75] | 0.369 |
| Oligodendrocytes | 17.2 [15.1;21.5] | 17.2 [16.9;17.5] | 1.000 |
| T-cell | 1.80 [1.65;1.85] | 1.70 [1.65;1.90] | 0.827 |
| VMC | 0.90 [0.70;1.00] | 1.10 [1.05;1.10] | 0.246 |
10.4 Pseudobulk
10.4.1 Microglia
| p_val | avg_log2FC | pct.1 | pct.2 | p_val_adj | |
|---|---|---|---|---|---|
| H2ac20 | 0.0e+00 | -0.1750223 | 1 | 1 | 0.0000036 |
| mt-Nd4l | 0.0e+00 | -0.4711669 | 1 | 1 | 0.0001628 |
| Tnfaip2 | 0.0e+00 | -0.4571713 | 1 | 1 | 0.0001815 |
| Gm13849 | 0.0e+00 | -0.0263631 | 1 | 1 | 0.0004293 |
| Prdm1 | 1.0e-07 | -0.1948147 | 1 | 1 | 0.0020465 |
| C430049B03Rik | 2.0e-07 | -0.1446463 | 1 | 1 | 0.0032855 |
| Man1a | 2.0e-07 | -0.2427612 | 1 | 1 | 0.0042481 |
| Ccl7 | 3.0e-07 | 0.0704861 | 1 | 1 | 0.0071494 |
| Il1a | 4.0e-07 | -0.5749078 | 1 | 1 | 0.0077532 |
| Cxcl1 | 6.0e-07 | -0.2383173 | 1 | 1 | 0.0136628 |
| Dusp2 | 1.1e-06 | -0.3441218 | 1 | 1 | 0.0242598 |
| Rpl35 | 1.4e-06 | -0.3722164 | 1 | 1 | 0.0304758 |
| Mir155hg | 1.5e-06 | -0.2601184 | 1 | 1 | 0.0321594 |
| mt-Atp8 | 1.9e-06 | -0.2756153 | 1 | 1 | 0.0409223 |
| Slc38a2 | 1.9e-06 | -0.1951182 | 1 | 1 | 0.0417004 |
Following treatment, microglia from the Alzheimer’s disease mouse model exhibited a broad transcriptional shift consistent with reduced pro-inflammatory activation and metabolic demand. Several mitochondrial genes (mt-Nd4l, mt-Atp8) and the amino acid transporter Slc38a2 were downregulated, suggesting diminished oxidative phosphorylation and nutrient uptake that normally support highly activated microglial states. Key inflammatory mediators, including Il1a, Cxcl1, Tnfaip2, and the microRNA host gene Mir155hg, also decreased, indicating attenuation of cytokine release, chemotactic signaling, NF-κB pathway activity, and miR-155–driven immune activation. Concomitant reductions in Prdm1 and regulatory non-coding transcripts (Gm13849, C430049B03Rik), together with decreased Rpl35 and Man1a, point to lower transcriptional activity, protein synthesis, and secretory function, while the decline in H2ac20 implies chromatin remodeling toward a less plastic state. Notably, this general dampening of inflammatory pathways was accompanied by increased expression of Ccl7, a chemokine involved in monocyte recruitment. This suggests that while most pro-inflammatory signals were suppressed, the treatment selectively maintained or enhanced pathways favoring controlled immune cell trafficking, potentially reflecting a rebalancing of microglial responses rather than their complete silencing. Collectively, these findings support the notion that the treatment shifts microglia away from a broadly activated phenotype toward a more homeostatic yet immunologically competent state, which may contribute to mitigation of chronic neuroinflammation in Alzheimer’s disease.
10.4.2 Astrocytes
| p_val | avg_log2FC | pct.1 | pct.2 | p_val_adj | |
|---|---|---|---|---|---|
| Wdr17 | 1.0e-07 | -0.6656742 | 1 | 1 | 0.0011904 |
| Lpar1 | 1.0e-07 | 0.5192552 | 1 | 1 | 0.0012595 |
| Dcc | 1.0e-07 | -0.6273549 | 1 | 1 | 0.0030505 |
| Wif1 | 7.0e-07 | 0.1848369 | 1 | 1 | 0.0159661 |
| Sorcs1 | 1.1e-06 | 0.3837130 | 1 | 1 | 0.0234440 |
| Tmsb4x | 1.8e-06 | 0.8196337 | 1 | 1 | 0.0383954 |
| Tenm2 | 2.2e-06 | 0.3594390 | 1 | 1 | 0.0476388 |
10.4.3 BAM
| p_val | avg_log2FC | pct.1 | pct.2 | p_val_adj | |
|---|---|---|---|---|---|
| Cxcl13 | 0e+00 | 1.9569000 | 1 | 0.333 | 0.0002040 |
| Wfdc17 | 2e-06 | -0.8501451 | 1 | 1.000 | 0.0435564 |
10.4.4 Choroid
| p_val | avg_log2FC | pct.1 | pct.2 | p_val_adj | |
|---|---|---|---|---|---|
| Rgs2 | 0 | 0.9370808 | 1 | 1 | 0.00068 |
10.4.5 Endothelial
| p_val | avg_log2FC | pct.1 | pct.2 | p_val_adj | |
|---|---|---|---|---|---|
| Gria1 | 1e-07 | 0.2876162 | 1 | 1 | 0.0014814 |
| Slc38a2 | 5e-07 | -0.3942290 | 1 | 1 | 0.0101714 |
10.4.6 Ependymal
| p_val | avg_log2FC | pct.1 | pct.2 | p_val_adj | |
|---|---|---|---|---|---|
| Mt3 | 1e-07 | -0.581966 | 1 | 1 | 0.0014678 |
10.4.7 Neurons
| p_val | avg_log2FC | pct.1 | pct.2 | p_val_adj | |
|---|---|---|---|---|---|
| Cldn5 | 0.0e+00 | 0.8740228 | 1.000 | 1.000 | 0.0000026 |
| Ly6c1 | 0.0e+00 | 0.8417682 | 1.000 | 1.000 | 0.0000039 |
| Hapln1 | 0.0e+00 | -1.0575334 | 0.667 | 1.000 | 0.0000206 |
| Slco1a4 | 0.0e+00 | 0.9098885 | 1.000 | 0.667 | 0.0000294 |
| Ptprb | 0.0e+00 | 0.7883225 | 1.000 | 0.667 | 0.0003976 |
| Vgf | 1.1e-06 | 0.8479086 | 1.000 | 1.000 | 0.0243259 |
| Ly6a | 1.6e-06 | 0.7068322 | 1.000 | 0.667 | 0.0353316 |
10.4.8 NPC
| p_val | avg_log2FC | pct.1 | pct.2 | p_val_adj | |
|---|---|---|---|---|---|
| Ikzf1 | 0.0e+00 | 1.0571115 | 1 | 1 | 0.0000004 |
| Runx1 | 0.0e+00 | 1.3537906 | 1 | 1 | 0.0000057 |
| Creg1 | 0.0e+00 | 1.0016099 | 1 | 1 | 0.0000301 |
| Gpr183 | 0.0e+00 | 1.1165005 | 1 | 1 | 0.0000624 |
| Adamts1 | 0.0e+00 | 0.9092272 | 1 | 1 | 0.0001537 |
| Inpp5d | 0.0e+00 | 1.1274030 | 1 | 1 | 0.0001906 |
| Zfp36 | 0.0e+00 | 1.0025476 | 1 | 1 | 0.0003610 |
| Lmo2 | 0.0e+00 | 0.9614975 | 1 | 1 | 0.0003979 |
| Lrmda | 0.0e+00 | 0.9927620 | 1 | 1 | 0.0009119 |
| Fth1 | 1.0e-07 | 0.7754402 | 1 | 1 | 0.0015168 |
| Sat1 | 1.0e-07 | 1.1250506 | 1 | 1 | 0.0016771 |
| Lpcat2 | 1.0e-07 | 0.9794508 | 1 | 1 | 0.0017612 |
| Unc93b1 | 1.0e-07 | 1.1107730 | 1 | 1 | 0.0018391 |
| Nfkbia | 1.0e-07 | 0.8980613 | 1 | 1 | 0.0019855 |
| Tmem119 | 1.0e-07 | 0.8763570 | 1 | 1 | 0.0020384 |
| Ptprc | 2.0e-07 | 0.9292133 | 1 | 1 | 0.0042320 |
| Lgals9 | 3.0e-07 | 0.7196859 | 1 | 1 | 0.0058081 |
| Adap2 | 3.0e-07 | 0.8139424 | 1 | 1 | 0.0071801 |
| P2ry12 | 6.0e-07 | 1.0277527 | 1 | 1 | 0.0118773 |
| Cd83 | 7.0e-07 | 0.9920499 | 1 | 1 | 0.0145304 |
| Itgam | 7.0e-07 | 1.0216380 | 1 | 1 | 0.0145724 |
| Abca9 | 8.0e-07 | 0.8031022 | 1 | 1 | 0.0174620 |
| Rnase4 | 8.0e-07 | 0.8590141 | 1 | 1 | 0.0179884 |
| Ly6e | 8.0e-07 | 0.8554275 | 1 | 1 | 0.0180234 |
| H2-T23 | 1.6e-06 | 0.7770506 | 1 | 1 | 0.0350168 |
| Selplg | 1.6e-06 | 0.7707752 | 1 | 1 | 0.0351740 |
| Lyn | 1.8e-06 | 0.9601579 | 1 | 1 | 0.0397079 |
| St3gal6 | 2.1e-06 | 0.8886279 | 1 | 1 | 0.0453242 |
10.4.9 Oligodendrocytes
| p_val | avg_log2FC | pct.1 | pct.2 | p_val_adj | |
|---|---|---|---|---|---|
| Ms4a6b | 5.0e-07 | 0.0686229 | 1 | 1 | 0.0096832 |
| C1qa | 8.0e-07 | 0.4272693 | 1 | 1 | 0.0167962 |
| Shoc2 | 9.0e-07 | -0.2455762 | 1 | 1 | 0.0196189 |
| Ctss | 1.2e-06 | 0.4387682 | 1 | 1 | 0.0254450 |
| mt-Atp8 | 1.3e-06 | -0.3642960 | 1 | 1 | 0.0273779 |
| mt-Nd4l | 2.0e-06 | -0.4666682 | 1 | 1 | 0.0426910 |
10.4.10 OPC
| p_val | avg_log2FC | pct.1 | pct.2 | p_val_adj | |
|---|---|---|---|---|---|
| Hpgd | 5e-07 | -1.446021 | 1 | 1 | 0.0109219 |
10.4.11 T-cell
| p_val | avg_log2FC | pct.1 | pct.2 | p_val_adj | |
|---|---|---|---|---|---|
| Dscam | 4e-07 | 0.8264654 | 1 | 1 | 0.008977 |
10.4.12 VMC
| p_val | avg_log2FC | pct.1 | pct.2 | p_val_adj | |
|---|---|---|---|---|---|
| Rbp4 | 0 | 1.454712 | 1 | 0.667 | 0.0003589 |
10.4.13 Number of DEGs up/down-regululated by aducanumab
